Decreased Collagen Deposition Research Articles

Human Allogeneic Liver-Derived Progenitor Cells Significantly Improve NAFLD Activity Score and Fibrosis in Late-Stage NASH Animal Model.

Accumulated experimental and clinical evidence supports the development of human allogeneic liver-derived progenitor cells (HALPCs) to treat fibro-inflammatory liver diseases. The aim of the present study was to evaluate their therapeutic effect in a non-alcoholic steatohepatitis (NASH)-STAM mouse model. The immune signaling characteristics of HALPCs were first assessed in vitro. Upon inflammation treatment, HALPCs secreted large amounts of potent bioactive prostaglandin E2 and indoleamine 2,3-dioxygenase, which significantly reduced CD4+ T-lymphocyte proliferation and secretion of proinflammatory cytokines. In vivo, HALPCs were intravenously administered as single or triple shots (of a dose of 12.5 × 106 cells/kg BW) in STAM mice. Transplantation of HALPCs was associated with a significant decrease in the NAFLD activity score at an early stage and in both inflammation and hepatocyte ballooning scores in late-stage NASH. Sirius red staining analyses revealed decreased collagen deposition in the pericentral region at both stages of NASH. Altogether, these findings showed the anti-inflammatory and anti-fibrotic features of HALPCs in an in vivo NASH model, which suggests their potential to reverse the progression of this chronic fibro-inflammatory disease.

Saponins from Panax japonicus alleviate adipose tissue fibrosis and metabolic dysfunction in high-fat-diet-induced obese mice

Introduction: Adipose tissue fibrosis is a typical feature of adipose tissue dysfunction in obese individuals, which is closely related to metabolic diseases. Objective: To explore the effect and mechanism of Saponins from Panax japonicus (SPJ) on adipose tissue fibrosis in obese mice induced by high fat diet (HFD). Materials and methods: We established a HFD induced obese mice model. Then the obese mice were treated with 90 mg/kg SPJ by oral gavage for 10 weeks. The levels of adipose tissue fibrosis and molecules related to signaling pathways were measured. Then the effects of SPJ on TGFβ1- induced fibrosis in 3T3-L1 differentiated adipocytes were evaluated. Results: SPJ reduced body weight, fat accumulation, and improved glucose and lipid metabolism in obese mice. SPJ decreased collagen deposition and expressions of fibrotic genes in epididymal white adipose tissue (eWAT) of obese mice. SPJ decreased the levels of TGFβ1 protein and pSmad2, and increased the expression of PPARγ and PGC1α, thus alleviating oxidative stress in eWAT. Consistently, SPJ inhibited TGFβ1- induced fibrosis in 3T3-L1 differentiated adipocytes. Conclusions: SPJ may alleviate adipose tissue fibrosis and improve obesity by inhibiting TGFβ1/Smad2 and activating PPARγ/PGC1α pathway. SPJ is expected to become an efficient medicine for treatment of obesity.

Molecular mechanisms of Agardhiella subulata attenuates hepatic fibrosis by modulating hepatic stellate cell activation via the reduction of autophagy

In this study, we investigated the effects of Agardhiella subulata extract (ASE) and its underlying mechanisms in vivo and in vitro. Rats induced hepatic fibrosis with CCl4 and transforming growth factor-β1 (TGF-β1) activated rat hepatic stellate cell line (HSC-T6) was used. The results showed that ASE (oral, 10 or 50 mg/kg) effectively attenuated liver fibrogenesis in the rats. This was evidenced by decreases in serum glutamic oxaloacetic transaminase and glutamic pyruvic transaminase levels, histopathological changes, decreased collagen deposition, and extracellular matrix formation. ASE significantly reduced hepatocyte apoptosis, Kupffer cell activation, and the levels of fibrogenic molecules such as alpha-smooth muscle actin and collagen. Furthermore, ASE downregulated the TGF-β1/Smad signaling pathway and concomitantly inhibited the activation of hepatic stellate cells by reducing autophagy and inducing the regeneration of lipid droplets both in vivo and in vitro. Overall, ASE inhibited various mechanisms that are important targets in the prevention of hepatic fibrosis.

CLEC5A is critical in Pseudomonas aeruginosa-induced NET formation and acute lung injury.

Pseudomonas aeruginosa is one of the most common nosocomial infections worldwide, and it frequently causes ventilator-associated acute pneumonia in immunocompromised patients. Abundant neutrophil extracellular traps (NETs) contribute to acute lung injury, thereby aggravating ventilator-induced lung damage. While pattern recognition receptors (PRRs) TLR4 and TLR5 are required for host defense against P. aeruginosa invasion, the PRR responsible for P. aeruginosa-induced NET formation, proinflammatory cytokine release, and acute lung injury remains unclear. We found that myeloid C-type lectin domain family 5 member A (CLEC5A) interacts with LPS of P. aeruginosa and is responsible for P. aeruginosa-induced NET formation and lung inflammation. P. aeruginosa activates CLEC5A to induce caspase-1-dependent NET formation, but it neither causes gasdermin D (GSDMD) cleavage nor contributes to P. aeruginosa-induced neutrophil death. Blockade of CLEC5A attenuates P. aeruginosa-induced NETosis and lung injury, and simultaneous administration of anti-CLEC5A mAb with ciprofloxacin increases survival rate and decreases collagen deposition in the lungs of mice challenged with a lethal dose of P. aeruginosa. Thus, CLEC5A is a promising therapeutic target to reduce ventilator-associated lung injury and fibrosis in P. aeruginosa-induced pneumonia.

Hesperetin mitigates sorafenib-induced cardiotoxicity in mice through inhibition of the TLR4/NLRP3 signaling pathway.

Sorafenib is an oral multi-kinase receptor inhibitor that targets various signaling pathways. It is used as the first line of treatment in advanced hepatocellular and renal cell carcinomas. Sorafenib was reported to induce cardiotoxicity due to myocyte necrosis. Hesperetin is a naturally occurring flavonoid with antioxidant and anti-inflammatory capabilities. This study investigated the putative protective effect of hesperetin against sorafenib-induced cardiotoxicity in mice through downregulation of NLRP3/TLR4 signaling and inhibition of apoptosis. Twenty-four male Swiss mice were distributed into four groups: untreated control, hesperetin (50 mg/kg/day, orally), sorafenib (100 mg/kg/day, orally), and combination (Hesperetin+Sorafenib). After a three-week treatment period, various biochemical parameters in cardiac tissues were assessed. TNF-α, IL-1β, and IL-6 levels were measured. Moreover, TLR4 and NLRP3 expressions were evaluated using Western blot analysis. Histopathological examination and immunohistochemical assessment of apoptotic activity were done. Compared with the sorafenib group, the combination group exhibited reduced TNF-α, IL-1β, IL-6 levels and lower NLRP3/TLR4 expressions. Histologically, the combination group showed improved myocardial histology and a marked decrease in collagen deposition. Immunohistochemical examination showed decreased caspase-3 and increased Bcl-2 expression. Before recommending hesperetin as an adjuvant, clinical studies are warranted for mitigating sorafenib cardiotoxicity.

Phloridzin Reveals New Treatment Strategies for Liver Fibrosis.

Liver fibrosis is an urgent public health problem which is difficult to resolve. However, various drugs for the treatment of liver fibrosis in clinical practice have their own problems during use. In this study, we used phloridzin to treat hepatic fibrosis in the CCl4-induced C57/BL6N mouse model, which was extracted from lychee core, a traditional Chinese medicine. The therapeutic effect was evaluated by biochemical index detections and ultrasound detection. Furthermore, in order to determine the mechanism of phloridzin in the treatment of liver fibrosis, we performed high-throughput sequencing of mRNA and lncRNA in different groups of liver tissues. The results showed that compared with the model group, the phloridzin-treated groups revealed a significant decrease in collagen deposition and decreased levels of serum alanine aminotransferase, aspartate aminotransferase, laminin, and hyaluronic acid. GO and KEGG pathway enrichment analysis of the differential mRNAs was performed and revealed that phloridzin mainly affects cell ferroptosis. Gene co-expression analysis showed that the target genes of lncRNA were obvious in cell components such as focal adhesions, intercellular adhesion, and cell–substrate junctions and in metabolic pathways such as carbon metabolism. These results showed that phloridizin can effectively treat liver fibrosis, and the mechanism may involve ferroptosis, carbon metabolism, and related changes in biomechanics.

Therapeutic induction of Bcl2-associated athanogene 3-mediated autophagy in idiopathic pulmonary fibrosis.

BackgroundExaggerated fibroblast proliferation is a well‐known feature in idiopathic pulmonary fibrosis (IPF) which may be – in part – due to insufficient autophagy, a lysosome dependent cellular surveillance pathway. Bcl2‐associated athanogene 3 (BAG3) is a pivotal co‐chaperone of the autophagy pathway. Here, we studied whether therapeutic modulation of BAG3‐mediated autophagy can rescue insufficient autophagy and impact IPF fibroblast proliferation.MethodsPrimary interstitial fibroblasts or precision cut lung slices (PCLS) of IPF lungs were treated with (1) the antifibrotic drug pirfenidone (Pirf), (2) the demethylating agent 5‐azacytidine (Aza), (3) the BAG3 modulator cantharidin (Ctd). Autophagy flux was measured following pretreatment with the autophagy inhibitors or by GFP‐RFP‐LC3B transfection followed by drug treatments. Proliferation was measured by 5‐bromo‐2′‐deoxyuridine assay. BAG3, filamin C (FLNC), proliferating‐cell‐nuclear‐antigen (PCNA), collagen1A1 (COL1A1) and autophagy proteins were assessed by immunoblotting or immunofluorescence. Loss of function experiments were performed by siRNA mediated knockdown of BAG3.ResultsIn comparison with healthy donors, increased BAG3 protein was observed in IPF lung homogenates and IPF fibroblasts. In addition, the substrate of BAG3‐mediated autophagy, FLNC, was increased in IPF fibroblasts, implying insufficient activation of BAG3‐dependent autophagy. Therapeutic modulation of this pathway using Aza and Ctd alone or in combination with the IPF therapy drug Pirf rescued the insufficient BAG3‐mediated autophagy and decreased fibroblast proliferation. Such effects were observed upon therapeutic modulation of BAG3 but not upon knock down of BAG3 per se in IPF fibroblasts. Similarly, PCLS of IPF patients showed a significant decrease in collagen deposition in response to these drugs, either alone or in a more potent form in combination with Pirf.ConclusionsOur study reveals that repurposing drugs that modulate autophagy regulating proteins render therapeutic benefits in IPF. Fine tuning of this pathway may hence signify a promising therapeutic strategy to ameliorate antifibrotic properties and augment the efficacy of current IPF therapy.

Extracts of Periplaneta americana alleviate hepatic fibrosis by affecting hepatic TGF-β and NF-κB expression in rats with pig serum-induced liver fibrosis.

Liver fibrosis is caused by continuous wound healing responses to various harmful stimuli, including viral infection, drugs, alcohol, and autoimmune liver disease. The purpose of this study was to examine the effects of extracts of Periplaneta americana (EPA) in rats with pig serum-induced liver fibrosis to preliminarily assess the antifibrotic effect of EPA. Seventy rats were randomly divided into 7 groups (10 rats in each group): HC, the healthy control group; FC, the fibrotic control group; TL, low-dose EPA treatment group group; TM, medium-dose EPA group; TH, high-dose EPA treatment group; TC1, Panax notoginseng/Salvia mitiorrhiza treatment control group 1; TC2, colchicine treatment control group 2. TC1 and TC2 were used as the positive control to demonstrate the difference between EPA and the effects of other compounds. The liver fibrosis model was induced by intraperitoneal injection of 0.5 mL pig serum twice a week for 13 weeks in all groups except for the HC group. The hepatic fibrosis model was established at the 7th week, and followingly, the corresponding compounds were administered once a day in all groups for 6 weeks. Alanine aminotransferase (ALT) and aspartate aminotransferase (AST) activity was determined in rat blood serum. We also measured liver fibrosis-related serum markers, including hyaluronic acid (HA), mucin layer (LN), type III pre-collagen (PC-III) and type IV collagen (IV-C). Hematoxylin and eosin (H&E) and Masson stainings were used to assess liver morphology and determine the stage of fibrosis. Immunohistochemistry was used to detect the protein expression of NF-κB, α-smooth muscle actin (α-SMA), transforming growth factor-β1 (TGF-β1) and tissue inhibitor of metalloproteinase-1 (TIMP-1) in rat liver tissue. Compared with that of the HC group, the liver tissue of the FC group presented obvious liver damage and collagen deposition. The serum levels of ALT, AST, HA, LN, PC-III and IV-C and the expression of NF-κB, α-SMA, TGF-β1 and TIMP-1 in the FC group were significantly higher than those in the HC group, the EPA treatment groups, the TC1 group and the TC2 group (P < 0.01). The levels of serum ALT, AST, HA, LN, PC-III and IV-C and the expression of α-SMA, NF-κB, TGF-β1 and TIMP-1 in the TL, TC1 and TC2 groups were significantly higher than those TM and TH groups (P < 0.05). EPA treatment significantly improved liver function, decreased collagen deposition and reversed the pathological changes related to liver fibrosis. We found that EPA could reduce liver inflammation, suppress liver cell degeneration and necrosis, and reduce the formation of liver fibrous tissue. Its mechanism might be associated with inhibiting the expression of TGF-β1, TIMP-1, NF-κB and α-SMA to block signal transduction pathways in the hepatic fibrosis process. Therefore, EPA, as a traditional Chinese medicine, might be potentially used to prevent and treat hepatic fibrosis in the future. However, further more experiments are necessary to verify its effectiveness and possible signaling pathways.

35. Human Adipose-derived Stem Cells Decrease Skin Damage in Mice After Scalp Irradiation: A Pilot Experiment

Purpose: The treatment for radiation dermatitis ulcers is surgical reconstruction. We aimed to characterize the effects of human adipose-derived stem cells (hAdMSCs) to prevent complications of radiation dermatitis of the scalp in an immunocompetent mouse model. Methods: Two 4-week-old female C57BL/6 mice were delivered a dose of 60 Gy to a skin area between the coronal and lambdoid sutures using a 5 mm circular collimator. On day 12 after irradiation, 600,000 luciferase- and green fluorescent protein-labeled hAdMSCs in 300 µL of PBS were injected subcutaneously into the scalp of one mouse. Mice were photographed under standardized conditions and weighted daily. Bioluminescence imaging (BLI) was performed daily from the day after injection until there was no detectable signal. Both were euthanized on day 27 after irradiation, and the scalp tissue was collected. Staining with hematoxylin and eosin, Masson’s trichrome, and immunohistochemistry for CD31 were performed. The histological slides were digitalized and analyzed using ImageJ (NIH). Statistical analyses were performed using Excel 2010 (Microsoft Corp., United States). Results: Bioluminescence images showed that hAdMSCs survived in vivo for five days. While desquamation and ulceration appeared on day 14 and lasted for 12 days in the treated mouse, the control mouse did not show signs of improvement by the time of euthanasia. The treated mouse had a greater mean erythematous area than the control mouse (0.38 cm2 vs. 0.23 cm2, p<0.05). The mean ulceration areas did not differ between the treated and control mouse (0.12 cm2 vs. 0.14 cm2, p>0.05). The epidermis, total skin thickness, and hypodermis of the irradiated regions were significantly thicker than the non-irradiated regions in both the treatment and control mice. The vessel size (treated = 67.25 µm2, control = 72.27 µm2, p>0.05) and the difference in tissue area occupied by vessels (treated = 2.58%, control = 2.42%, p>0.05) of the radiated regions did not differ between mice. Collagen deposition in the irradiated region was decreased in the treated mouse compared to the control mouse (11% vs. 42%). Conclusion: Human adipose-derived stem cells decrease the duration of ulceration and improve the macroscopic quality of the skin, survive for only 5 days after transplantation, decrease collagen deposition in the irradiated area, increase micro vessel area and size, and decrease the thickness of the irradiated area of the skin compared to controls two weeks after transplantation.

68. Adipocytes Transition to Pro-fibrotic Fibroblasts and Contribute to Muscle Fibrosis Following Nerve Injury

Purpose: Recent studies have suggested that interconversion between adipocytes and fibroblasts (the cellular mediators of skin scarring) occurs in the wound environment. Muscle fibrosis occurs following aging, muscle injury, and muscular dystrophies causing weakness and pain. The role of adipocytes in muscle fibrosis has not been fully explored. Our study examines whether mature adipocytes participate in muscle fibrosis following nerve injury via conversion into dermal fibroblasts. Methods:AdipoqCre transgenic driver mice were crossed to R26mTmG reporter mice to generate AdipoqCre;ROSA26mTmG mice, in which mature adipocytes (Adipoq/adiponectin-expressing cells) express green fluorescent protein (GFP), and all other cells express a red fluorescent protein (RFP). AdipoqCre;ROSA26mTmG mice underwent sciatic nerve injury per established protocol. Injured tibialis anterior and gluteus maximus muscle was harvested at postoperative day-14 (POD14) and underwent immunofluorescent staining for fibroblast (collagen type IV/col-IV, alpha-smooth muscle actin/a-SMA) and adipocyte (perilipin) cell markers. To achieve local adipocyte ablation, AdipoqCre;ROSA26mTmG;R26tm1(HBEGF)Awai mice were generated and wounded, and diphtheria toxin (DT;400ng/mouse) was injected into the muscle at the nerve injury site. DT- and vehicle control-treated muscle underwent histologic analysis. Results: We identified a significantly greater number of adiponectin-lineage-positive cells (GFP+) within injured muscle at POD14 compared to uninjured muscle (*p<0.05,n=5). The GFP+ cells that infiltrated the muscle injury exhibited upregulation of fibrotic markers including col-IV and α-SMA, and downregulation of adipocyte markers including perilipin, indicating conversion of adipocytes into pro-fibrotic fibroblasts. DT-induced ablation of Adipoq-lineage-positive cells resulted in reduced muscle scarring, with decreased collagen deposition compared to vehicle-control injured muscle (*p<0.05,n=5). Conclusion: Our study demonstrates that adipocytes undergo conversion to pro-fibrotic fibroblasts in response to nerve injury and contribute to muscle fibrosis. Targeted adipocyte ablation resulted in reduced muscle scarring, suggesting that adipocyte-derived fibroblasts are important contributors to muscle fibrosis.

Mebendazole, an anti-helminth drug, suppresses inflammation, oxidative stress and injury in a mouse model of ulcerative colitis

Mebendazole (MBZ) is an efficacious anthelmintic with known anti-inflammatory and fibrinolytic properties. In this study, we aimed to explore the protective effects of this FDA-approved drug against DSS-induced colitis in a murine model either alone or in combination with Sulfasalazine (SSZ), a standard therapy for ulcerative colitis. We found that MBZ significantly improved colitis disease activity index as assessed by changes in body weight, degree of stool consistency, rectal bleeding, and prolapse. We also found that MBZ ameliorated the colon histopathological score by attenuating crypt loss, mucosal damage, and inflammation score in colitis tissues. Similarly, DSS-induced colon shortening, colon weight loss, and increase in spleen weight were all abrogated in the presence of MBZ. Moreover, MBZ decreased inflammation, possibly by reducing oxidative stress markers, suppressing inflammatory cell infiltration, and down-regulation of inflammatory genes in colon tissues. Furthermore, MBZ potently reduced fibrosis by decreasing collagen deposition and down-regulating pro-fibrotic genes including Col 1a1 and Col 1a2 in colitis tissue homogenates. In conclusion, our study showed that this broad-spectrum anthelminthic could be repurposed as a novel therapy for ulcerative colitis without any observed side effects, however, regarding the concerns about the potential toxicity of MBZ in UC patients, future experiments on MBZ therapy in other models of UC is needed to completely address the toxicity concerns.

Application of Collagen-Based Scaffolds for the Treatment of Spinal Cord Injuries in Animal Models: A Literature Update

SCI is regarded as one of the most devastating central nervous system (CNS) injuries, exhibiting an alarmingly rising incidence rate, indirectly connected with the expansion of the global economy. The consequences of SCI are multidimensional: SCI injuries may result in permanent voluntary motor dysfunction and loss of sensation while incurring heavy economic and psychological burdens as part of the treatment. Thus, it is crucial to develop effective and suitable SCI treatment strategies. Collagen-based scaffold application is one of the most promising methods of SCI treatment.This review compiles newer bibliographical data regarding the application of collagen scaffolds for the treatment of Spinal cord injury (SCI) in animal models. Recently, several relevant studies have been carried out using carefully selected animals with similar pathophysiology to humans. In mouse, rat and canine models that have undergone transection or hemisection, the stump connection, the transplanted cell differentiation, and the elimination of glial scar are promising. Also, encouraging results have been found regarding the increased neuronal growth, the decreased collagen deposition, the behavioral recovery, the improved electrophysiology, and the enhanced axonal regeneration.

Abstract 3191: Tumor intrinsic HuR promotes stroma activation in pancreatic cancer

Abstract Background: One of the factors contributing to poor prognosis of pancreatic ductal adenocarcinoma (PDAC) is thought to be the characteristically dense stroma and heterogeneous tumor microenvironment. The PDAC stroma involves multiple players, including cancer associated fibroblasts (CAFs) and pancreatic stellate cells (PSCs). These stromal cells, when activated, promote extensive fibrosis in PDAC, which is associated with poor prognosis and chemoresistance. We have previously established that the RNA-binding protein Human Antigen R (HuR, ELAVL1) facilitates PDAC progression. In a tumor microarray of matched normal and malignant patient tissues, we found high active (i.e. cytoplasmic) HuR staining in 80% of PDAC tumors, while low or absent in adjacent normal tissues (n=80). Moreover, in both the orthotopic and the genetically engineered mouse models of PDAC, active HuR staining in tumors was dramatically higher than in normal pancreas. Pancreas-specific overexpression of HuR in a transgenic mouse model did not initiate tumorigenesis, but did increase fibrosis compared to control mice. We hypothesize that tumor intrinsic HuR regulates transcripts that are needed for tumor cells to activate the surrounding stroma. Methods: MiaPaCa-2 or PANC-1 human PDAC cells, either HuR-proficient or CRISPR-mediated knocked-out, were used in an orthotopic model where cells were injected into the tail of the pancreas. Both NRG and athymic mice were used. Resulting tumors underwent immunohistochemistry, immunofluorescence, or RNA-sequencing. Secreted proteins from cells in 2D culture were analyzed using a cytokine array. Results: HuR knockout tumors showed a decrease in CAF and PSC markers (i.e. podoplanin, alpha-SMA, desmin; &amp;gt;55% decrease, p&amp;lt;0.05), as well as a decrease in collagen deposition (56% decrease, p=0.0004). Additionally, HuR expression in tumors positively correlated with collagen abundance (p= 0.0002). RNA sequencing of PDAC tumors at various times during tumor development found that gene expression of stroma-activating ligands was significantly downregulated in HuR knockout tumors (e.g. TGFb, CCL2, PDGFA; -0.8&amp;lt;log2FC&amp;lt;-3.6, 0.0009&amp;lt;padj&amp;lt;2^-57). Moreover, we found a significant decrease of these stroma-activating ligands in conditioned media obtained from HuR knockout compared to wild type (e.g. PDGFAA, PTX3, CCL2; &amp;gt;50% decrease, p&amp;lt;0.0001). Conclusions: We found inhibition of HuR led to a robust decrease in stroma activation (i.e. lack of collagen, CAFs, and PSCs). To our knowledge, this is the first report of PDAC tumor intrinsic HuR having an extrinsic effect on neighboring stromal cells. Specifically, we found loss of HuR results in a decrease of secreted stroma-activating ligands, likely to be the cause of the decreased fibrosis in HuR knockout tumors. With evidence that fibrosis is associated with poor survival rates and chemoresistance, inhibiting HuR may sensitize tumors to standard of care therapy. Citation Format: Grace A. McCarthy, Jennifer M. Finan, Aditi Jain, Roberto DiNiro, Aaron Grossberg, Jonathan R. Brody. Tumor intrinsic HuR promotes stroma activation in pancreatic cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3191.

F4/80hi Resident Macrophages Contribute to Cisplatin-Induced Renal Fibrosis.

Cisplatin-induced kidney injury remains a major obstacle in utilizing cisplatin as a chemotherapeutic for solid-organ cancers. Thirty percent of patients treated with cisplatin develop acute kidney injury (AKI), and even patients who do not develop AKI are at risk for long-term declines in kidney function and development of chronic kidney disease (CKD). Modeling cisplatin-induced kidney injury in mice has revealed that repeated low doses of cisplatin lead to development of kidney fibrosis. This model can be used to examine AKI-to-CKD transition processes. Macrophages play a role in some of these processes, including immune response, wound healing, and tissue remodeling. Depleting macrophage populations in the kidney reduced fibrosis development in other models of renal fibrosis. We used either C57BL/6 mice with a Ccr2 genetic knockout or liposome encapsulated clodronate (Clodrosome) to deplete macrophage populations during repeated 9 mg/kg cisplatin treatments. We assessed how immune cell populations were altered in the blood and kidney of these mice and how these alterations affected development of renal fibrosis and kidney injury. We found that Clodrosome treatment decreased collagen deposition, myofibroblast accumulation, and inflammatory cytokine production, whereas Ccr2 genetic knockout had no effect on these markers after cisplatin treatment. Additionally, Ccr2-/- mice had decreased levels of F4/80lo infiltrating macrophages in the kidney after cisplatin treatments, but Clodrosome treatment depleted F4/80hi resident and CD206+ M2 macrophages. These data suggest that Clodrosome depletion of F4/80hi and M2 macrophages in the kidney attenuates development of renal fibrosis after repeated low doses of cisplatin.

Revealing the mechanism of Jiegeng decoction attenuates bleomycin-induced pulmonary fibrosis via PI3K/Akt signaling pathway based on lipidomics and transcriptomics

BackgroundPulmonary fibrosis (PF) is a serious lung disease with unknown etiology and irreversible course. Jiegeng decoction (JGD), a traditional prescription, is widely used to treat lung diseases due to its anti-inflammatory and expectorant effects. PurposeTo explore the effect of JGD on mice with PF and its underlying mechanism. For this purpose, we established a mouse model with PF by bleomycin (BLM) and then administered JGD and pirfenidone at different concentrations. ResultsIn vivo, JGD was found to reduce lung inflammation, improve lung function and decrease collagen deposition to alleviate bleomycin-induced PF in mice. The mouse lung tissue was analyzed using lipidomics and transcriptomics. We found phosphatidylinositol was decreased after JGD treatment in lipidomics results, while transcriptomics results showed the critical roles of PI3K/Akt signaling pathway in JGD treatment group. Then, Western Blot and Immunohistochemistry were used to validate that JGD may regulate the expression of Bax, Caspase3, Caspase8, Caspase9 and Bcl-2 apoptosis-related proteins via PI3K/Akt signaling pathway. TUNEL staining revealed that apoptosis mainly occurs on AEC IIs. ConclusionOur results showed that JGD inhibits apoptosis through the PI3K/Akt signaling pathway, thereby protecting against BLM-induced PF. Hence, JGD is expected to be a potential drug candidate for the treatment of PF.

The transforming growth factor beta (TGF-β/Smads) pathway regulates collagen synthesis and deposition in swim bladder of Chu's croaker (Nibea coibor) stimulated by proline

The swim bladder performs various functions in fish, is consumed by humans and can be processed into high-grade fish glue (traditional high-value tonic). However, little is known on the optimum amount of proline (Pro) required for promoting collagen synthesis and deposition in fish swim bladder and the regulatory mechanism. This study explored the effects and optimum amount of dietary Pro supplementation on collagen synthesis and deposition in Chu's croaker (Nibea coibor) swim bladder and its possible molecular mechanism. A total of 450 juvenile fish (8.64 ± 0.14 g) were randomly stocked into 18 cages (25 fish per cage) in triplicate and fed Pro supplemented diets (0, 5, 10, 15, 20 and 25 g kg−1) for eight weeks. Another 225 fish (11.62 ± 0.15 g) were distributed randomly into control, Pro and Pro diet plus injected with specific inhibitor of Smad3 (SIS3) in order to inhibit the transforming growth factor beta (TGF-β)/Smads pathway. Pro supplementation up-regulated significantly the expression of genes involved in TGF-β/Smads pathway such as collagen, type I, alpha 1 (col1a1) and 2 (col1a2), TGF-β and Smad2/3, and increased collagen content in the Chu's croaker swim bladder (P < 0.05). The optimum dietary Pro supplementation for deposition of collagen in Chu's croaker swim bladder was 19.36 g kg−1. The collagen content in the swim bladder positively correlated significantly with the mRNA expression of col1a1, col1a2, TGF-β and Smad2/3 genes (P < 0.05). The optimum dietary Pro supplementation for maximum synthesis of collagen in Chu's croaker swim bladder was 13.25, 13.32 and 15.94 g kg−1 based on mRNA expression of Smads2, Smads3 and col1a2 genes, respectively. SIS3 down-regulated the expression of col1a1, col1a2, TGF-β and Smad2/3 genes, subsequently decreased collagen deposition in the swim bladder (P < 0.05). These results suggest that 13.25 to 19.36 g kg−1 Pro supplementation improve collagen synthesis and deposition in Chu's croaker swim bladder. The TGF-β/Smad signaling pathway regulates collagen synthesis and deposition in Chu's croaker swim bladder. Our results provide an understanding on the molecular mechanism of collagen synthesis and deposition in fish for producing collagen required for human food and fish glue.

D-Carvone Attenuates CCl4-Induced Liver Fibrosis in Rats by Inhibiting Oxidative Stress and TGF-ß 1/SMAD3 Signaling Pathway.

Simple SummaryLiver fibrosis is a challenging global health problem resulting in a significant morbidity and mortality rates worldwide due to its rapid progression to cirrhosis and hepatocellular carcinoma. Therefore, identifying nontoxic therapies with precise curative effects to slow the progression of liver fibrosis comprises one of the most popular and high-priority areas of current research. D-carvone is a naturally occurring monoterpene abundant in the essential oil of aromatic plants such as caraway and spearmint. In the present study, the protective impact of D-carvone on carbon tetrachloride (CCl4)-induced liver fibrosis in rats was evaluated. Administration of D-carvone significantly enhanced liver functions, oxidant/antioxidant balance as well as liver histology. D-carvone ameliorated the progression of liver fibrosis, evident by the decreased collagen deposition (fibrosis score) and the reduced expression of the pro-fibrogenic markers TGF-β1 and SMAD3 in the liver. These findings reveal the anti-fibrotic effects of D-carvone and suggest that D-carvone could be a promising candidate for therapeutic intervention of liver fibrosis and other oxidative stress-related hepatic diseases.D-carvone is a natural monoterpene found in abundance in the essential oil of aromatic medicinal plants with a wide range of pharmacological values. However, the impact of D-carvone on liver fibrosis remains unclear. This study aimed to evaluate the anti-fibrotic potential of D-carvone in a rat model of liver fibrosis and to clarify the possible underlying mechanisms. Liver fibrosis was induced in rats by carbon tetrachloride, CCl4 (2.5 mL/kg, interperitoneally every 72 h for 8 weeks). Oral treatment of rats with D-carvone (50 mg/kg, daily) started on the 3rd week of CCl4 administration. D-carvone significantly enhanced liver functions (ALT, AST), oxidant/antioxidant status (MDA, SOD, GSH, total antioxidant capacity; TAC), as well as histopathological changes. Moreover, D-carvone effectively attenuated the progression of liver fibrosis, evident by the decreased collagen deposition and fibrosis score by Masson trichrome staining (MT) and α-SMA protein expression. Moreover, D-carvone administration resulted in a significant downregulation of the pro-fibrogenic markers TGF-β1 and SMAD3 and upregulation of MMP9. These findings reveal the anti-fibrotic effect of D-carvone and suggest regulation of the TGF-β1/SMAD3 pathway, together with the antioxidant activity as a mechanistic cassette, underlines this effect. Therefore, D-carvone could be a viable candidate for inhibiting liver fibrosis and other oxidative stress-related hepatic diseases. Clinical studies to support our hypothesis are warranted.

Target Discovery Platform Identifies Cardioprotective Genes Using Hibernation Omics Data

Over the course of their evolution, many mammals have naturally solved a number of human biomedical problems. The genes underlying their protective strategies are likely neither novel nor species‐specific, but rather shared among all mammals, including humans. Identifying and validating these genes could therefore lead to new therapeutic treatments for human clinical indications. To this end, we developed a target discovery platform which mines multi‐omics data from mammals resistant to human pathologies in order to identify novel therapeutic genetic targets. More specifically, our platform is built upon transcriptomic and epigenomic data from bio‐banked 13‐lined ground squirrel (13‐LGS) tissues collected at precise seasonal and physiological time‐points across their annual cycle of hibernation. Our platform also incorporates myriad genomic, transcriptomic, and proteomic datasets publicly available for this and other disease‐resistant species, as well as human genomics and transcriptomics data. Here we sought to identify novel genetic targets protective against cardiac fibrosis. While the hearts of 13‐LGS experience repeated bouts of ischemia‐reperfusion during hibernation, long‐term fibrotic scarring resulting from this type of injury is not detected. To identify protective targets, we first generated RNASeq data from hearts of 13‐LGS across 11 different seasonal and physiological groups (n≥5 per group). After obtaining transcript abundance counts for each sample library, we performed weighted correlation gene network analysis (WGCNA) to identify modules of transcripts co‐expressed among the different physiological groups. One resulting WGCNA module contained TGFB1 (transforming growth factor beta 1), a known central mediator of fibrogenesis. Knockdown (KD) of this module’s hub gene, designated here as FAUN211G, as well as the second top‐ranking gene, designated FAUNA212G, significantly decreased collagen deposition in an in vitro 2D model of cardiac fibrosis using primary human cardiac fibroblasts. We next perturbed these genes in an in vitro 3D spheroid model of human cardiac fibrosis and detected improved parameters of cardiac function. Finally, we tested small molecule inhibitors on FAUNA211G in both 2D and 3D in vitro models; we identified significant reductions in myofibroblast activation, proliferation and extracellular matrix deposition. Both FAUNA211G and FAUNA212G were previously unknown to be involved in cardiac fibrosis. In sum, we demonstrate that network analysis of the hibernator’s transcriptome is a strategy to identify novel target genes for treatment of human clinical indications.

Aktivitas Antibakteri Ekstrak Bawang Hitam Variasi Waktu Aging Terhadap Pencegahan Dysbiosis Kulit Penyebab Jerawat

Skin diseases, including acne, can be caused by a microbial imbalance (dysbiosis). The specific bacteria involved in acne dysbiosis are Cutibacterium (Propionibacterium) acnes and Staphylococcus epidermidis. Black garlic has better potential to be alternative antibiotic and chemical to curing ance and maintaining skin health, because it has antimicrobial potential and can inhibit the decrease in collagen deposition on the skin. These potentials can be alternative of antibiotics and chemicals to treat acne and maintain healthy skin. This study aimed to determine the antibacterial activity of black garlic extract against bacteria that cause acne dysbiosis. Garlic was aging with 3 variations of time (7, 14, and 21 days) then macerated with 70% alcohol and tested for antimicrobial activity to Cutibacterium acnes and Staphylococcus epidermidis with clindamycin as positive control. Black garlic extract provided inhibitory activity against Staphylococcus epidermidis and Cutibacterium acnes. The best recommendation time for aging garlic to prevent skin dysbiosis causes acne is 7 to 14 days.

Composite Fish Collagen-Hyaluronate Based Lyophilized Scaffolds Modified with Sodium Alginate for Potential Treatment of Chronic Wounds.

Chronic wounds are characterized by both decreased collagen deposition and increased collagen breakdown. It is reasonable to hypothesize that exogenous collagen can potentially promote wound healing by reducing degradation enzymes in the wound environment and disrupting the cycle of chronicity. Therefore, this study aimed to develop an optimal combination of fish collagen (FCOL), sodium alginate (SA), and hyaluronic acid (HA) loaded with bovine serum albumin (BSA) as a model protein fabricated as lyophilized scaffolds. The effects of sodium alginate (SA#) with higher mannuronic acid (M) were compared to sodium alginate (SA*) with higher guluronic acid (G). The SA* with higher G resulted in elegant scaffolds with hardness ranging from 3.74 N–4.29 N that were able to withstand the external force due to the glycosidic bonds in guluronic acid. Furthermore, the high G content also had a significant effect on the pore size, pore shape, and porosity. The water absorption (WA) ranged from 380–1382 (%) and equilibrium water content (EWC) 79–94 (%) after 24 h incubation at 37 °C. The SA* did not affect the water vapor transmission rate (WVTR) but incorporating BSA significantly increased the WVTR making these wound dressing scaffolds capable of absorbing about 50% exudate from a heavily exuding chronic wound. The protein released from the composite systems was best explained by the Korsmeyer–Peppas model with regression R2 values ranging from 0.896 to 0.971 and slope or n < 0.5 indicating that the BSA release mechanism was governed by quasi-Fickian diffusion. Cell viability assay showed that the scaffolds did not inhibit the proliferation of human dermal fibroblasts and human epidermal keratinocytes, and are therefore biocompatible. In vitro blood analysis using human whole blood confirmed that the BSA-loaded SA*:FCOL:HA scaffolds reduced the blood clotting index (BCI) by up to 20% compared to a commercially available sponge for chronic wounds. These features confirm that SA*:FCOL:HA scaffolds could be applied as a multifunctional wound dressing.